Extracts from garlic dry powders modulate LPS-induced liberation of cytokines. H.-P. Keiß 1 , V.M. Dirsch 1 , T. Hartung

1. DMSO extracts Il-1 TNF- Aqueous extracts Il-1 TNF- 3. NF-B activity assay To investigate whether the modulation of cytokine levels in human whole blood leads to a change in the transactivation activity of the transcription factor NF- B we performed a NF- B-luciferase reporter gene assay. Therefore cells were transfected as described under methods and subsequently stimulated with the supernatants of human blood, treated with DMSO garlic extracts and LPS or with LPS alone. Aqueous extracts DMSO extracts Il-10 NF- B activity assay unfertilized garlic fertilized garlic Extracts from garlic dry powders modulate LPS-induced liberation of cytokines. H.-P. Keiß1, V.M. Dirsch1, T. Hartung2, A.M. Vollmar1 1Department of Pharmacy, Center of Drug Research, Butenandtstr 5-13, 81377 Munich, Germany 2Biochemical Pharmacology, University of Konstanz, Universitaetsstr. 10, 78457 Konstanz, Germany Hans.Keiss@cup.uni-muenchen.de Methods: Garlic extracts Garlic dry powder from unfertilized or sulphur fertilized allium sativum L. cv. Printanor was used. Garlic dry powders were extracted either with aqueous media or DMSO. Cytokine liberation assay Heparinized human whole blood was exposed to respective garlic extracts and stimulated with lipopolysacharide (LPS) 10 ng/ml. Cytokines in the cell free supernatants were collected and subsequently quantified by sandwich ELISA. NF-B luciferase reporter gene assay HEK 293 cells were transfected with a plasmid containing a NF- B driven firefly-luciferase gene and a plasmid containing a HSV-TK driven renilla-luciferase gene. Cells were stimulated with supernatants from the cytokine liberation assay and NF-B promoter activity was measured with a Dual-Luciferase Reporter System. Background: Garlic (Allium sativum) has been proposed to have anti-atherosclerotic potential, with sulphur compounds seemingly the pharmacologically active substances.Increasing evidence is given that inflammatory processes, such as NF-B activation, cytokine liberation or expression of adhesion molecules, are pivotal in the pathophysiology of atherosclerosis. Aim: The aim of our study was to examine the effects of different garlic extracts on cytokine production and to elucidate the impact of increasing amounts of sulphur compounds on the biological activity of the extracts. 1. Liberation of pro-inflammatory cytokines • DMSO extracts do not liberate Il-1 and TNF- • DMSO extracts reduce LPS-induced liberation of Il-1 and TNF- • Fertilization has no influence on the inhibitory effect • Aqueous extracts liberate Il-1 and TNF- • Aqueous garlic extracts reduce LPS-induced liberation of Il-1 and TNF-, however not dose dependently. • Fertilized garlic shows a weaker pro-inflammatory, but a stronger anti- inflammatory potential than unfertilized garlic 2. Liberation of anti-inflammatory cytokine • Aqueous garlic extracts liberate Il-10 • Aqueous extracts increase LPS-induced liberation of Il-10 • Low concentrations of DMSO garlic extracts increase LPS-induced liberation of Il-10 • High concentrations of aqueos fertilized garlic extract show a strong inhibitory effect on Il-10 liberation • Supernatants of blood cells treated with DMSO garlic extracts reduce NF-B activity • Fertilized garlic was more active than unfertilized garlic * < 0.05 Anova with Bonferroni post-test ** < 0.01 Anova with Bonferroni post-test Conclusion: This study indicates, that garlic dry powder extracts are able to decrease LPS-induced liberation of pro-inflammatory cytokines Il-1b and TNF-a. On the other hand, they can increase the liberation of the anti-inflammatory cytokine Il-10. These effects are strongly dependent on the solvent used for extraction. Furthermore, it was shown that sulphur fertilization may have a positive influence on the action mediated by garlic. In summary, DMSO garlic extracts modify cytokine release from human blood in a way possibly resulting in a lower expression of NF- B regulated genes. This work was supported by the EU : “Garlic and Health Project”, Id-Nr:QLK1-CT-1999-00498